Overview

  • Product name
    Anti-ErbB 2 antibody [EPR19547-12]
    See all ErbB 2 primary antibodies
  • Description
    Rabbit monoclonal [EPR19547-12] to ErbB 2
  • Host species
    Rabbit
  • Specificity
    Only reacts with mouse weakly in 4T1 cells
  • Tested applications
    Suitable for: ICC/IF, IP, Flow Cyt, IHC-P, WBmore details
  • Species reactivity
    Reacts with: Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human ErbB 2 aa 1200 to the C-terminus (phospho Y1248). The exact sequence is proprietary. (ab214275 recognizes both phospho and non-phospho ErbB2).
    Database link: P04626

  • Positive control
    • WB: Human ErbB 2 fragment recombinant protein; human breast cancer lysate; HeLa, A431, SK-BR-3, C6 and PC-12 whole cell lysates. IHC-P: Human breast cancer tissue. ICC/IF: SK-BR-3 cells. Flow Cyt: SK-BR-3 cells. IP: HeLa whole cell lysate.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab214275 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ICC/IF 1/250.
IP 1/30.
Flow Cyt 1/60.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

IHC-P 1/4000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB 1/1000. Detects a band of approximately 180 kDa (predicted molecular weight: 137 kDa).

Target

  • Function
    Protein tyrosine kinase that is part of several cell surface receptor complexes, but that apparently needs a coreceptor for ligand binding. Essential component of a neuregulin-receptor complex, although neuregulins do not interact with it alone. GP30 is a potential ligand for this receptor. Regulates outgrowth and stabilization of peripheral microtubules (MTs). Upon ERBB2 activation, the MEMO1-RHOA-DIAPH1 signaling pathway elicits the phosphorylation and thus the inhibition of GSK3B at cell membrane. This prevents the phosphorylation of APC and CLASP2, allowing its association with the cell membrane. In turn, membrane-bound APC allows the localization of MACF1 to the cell membrane, which is required for microtubule capture and stabilization.
    In the nucleus is involved in transcriptional regulation. Associates with the 5'-TCAAATTC-3' sequence in the PTGS2/COX-2 promoter and activates its transcription. Implicated in transcriptional activation of CDKN1A; the function involves STAT3 and SRC. Involved in the transcription of rRNA genes by RNA Pol I and enhances protein synthesis and cell growth.
  • Tissue specificity
    Expressed in a variety of tumor tissues including primary breast tumors and tumors from small bowel, esophagus, kidney and mouth.
  • Involvement in disease
    Hereditary diffuse gastric cancer
    Glioma
    Ovarian cancer
    Lung cancer
    Gastric cancer
    Chromosomal aberrations involving ERBB2 may be a cause gastric cancer. Deletions within 17q12 region producing fusion transcripts with CDK12, leading to CDK12-ERBB2 fusion leading to truncated CDK12 protein not in-frame with ERBB2.
  • Sequence similarities
    Belongs to the protein kinase superfamily. Tyr protein kinase family. EGF receptor subfamily.
    Contains 1 protein kinase domain.
  • Post-translational
    modifications
    Autophosphorylated. Autophosphorylation occurs in trans, i.e. one subunit of the dimeric receptor phosphorylates tyrosine residues on the other subunit (Probable). Ligand-binding increases phosphorylation on tyrosine residues (PubMed:27134172). Signaling via SEMA4C promotes phosphorylation at Tyr-1248 (PubMed:17554007). Dephosphorylated by PTPN12 (PubMed:27134172).
  • Cellular localization
    Cytoplasm. Nucleus and Cell membrane. Cytoplasm, perinuclear region. Nucleus. Translocation to the nucleus requires endocytosis, probably endosomal sorting and is mediated by importin beta-1/KPNB1.
  • Information by UniProt
  • Database links
  • Alternative names
    • Verb b2 erythroblastic leukemia viral oncogene homolog 2, neuro/glioblastoma derived oncogene homolog antibody
    • C erb B2/neu protein antibody
    • CD340 antibody
    • CD340 antigen antibody
    • Cerb B2/neu protein antibody
    • CerbB2 antibody
    • Erb b2 receptor tyrosine kinase 2 antibody
    • ERBB2 antibody
    • ERBB2_HUMAN antibody
    • HER 2 antibody
    • HER 2/NEU antibody
    • HER2 antibody
    • Herstatin antibody
    • Human epidermal growth factor receptor 2 antibody
    • Metastatic lymph node gene 19 protein antibody
    • MLN 19 antibody
    • MLN19 antibody
    • NEU antibody
    • NEU proto oncogene antibody
    • Neuro/glioblastoma derived oncogene homolog antibody
    • Neuroblastoma/glioblastoma derived oncogene homolog antibody
    • NGL antibody
    • p185erbB2 antibody
    • Proto-oncogene c-ErbB-2 antibody
    • Proto-oncogene Neu antibody
    • Receptor tyrosine-protein kinase erbB-2 antibody
    • TKR1 antibody
    • Tyrosine kinase type cell surface receptor HER2 antibody
    • Tyrosine kinase-type cell surface receptor HER2 antibody
    • V erb b2 avian erythroblastic leukemia viral oncogene homolog 2 (neuro/glioblastoma derived oncogene homolog) antibody
    • V erb b2 avian erythroblastic leukemia viral oncogene homolog 2 antibody
    • V erb b2 avian erythroblastic leukemia viral oncoprotein 2 antibody
    • V erb b2 erythroblastic leukemia viral oncogene homolog 2, neuro/glioblastoma derived oncogene homolog (avian) antibody
    • V erb b2 erythroblastic leukemia viral oncogene homolog 2, neuro/glioblastoma derived oncogene homolog antibody
    • Verb b2 erythroblastic leukemia viral oncogene homolog 2, neuro/glioblastoma derived oncogene homolog (avian) antibody
    see all

Images

  • All lanes : Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/1000 dilution

    Lane 1 : 4T1 (Mouse mammary gland carcinoma epithelial cell) whole cell lysate
    Lane 2 : 4T1(Mouse mammary gland carcinoma epithelial cell) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

    Predicted band size: 137 kDa
    Observed band size: 180 kDa
    why is the actual band size different from the predicted?



    Blocking and diluting buffer: 5% NFDM/TBST. 

  • Immunofluorescent analysis of 100% methanol-fixed SK-BR-3 (Human mammary gland adenocarcinoma cell line) cells labeling ErbB 2 with ab214275 at 1/250 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing membrane staining on SK-BR-3 cells.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)  (red).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Immunohistochemical analysis of paraffin-embedded Human breast cancer tissue labeling ErbB 2 with ab214275 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Membrane staining on Human breast cancer is observed [PMID: 18437174].

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

  • All lanes : Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/1000 dilution

    Lane 1 : Human ErbB 2 fragment recombinant protein
    Lane 2 : Human ErbB 4 fragment recombinant protein

    Lysates/proteins at 0.02 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 137 kDa


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    Human ErbB 2 fragment recombinant protein contains aa955-1255 with a His-Tag®; Human ErbB 4 fragment recombinant protein contains aa1000-1300 with a His-Tag®.

  • Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/2000 dilution + Human breast cancer lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

    Predicted band size: 137 kDa
    Observed band size: 180 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The molecular weight is consist with literature PMID :15741050.

  • Lanes 1-2 : Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/1000 dilution
    Lane 3 : Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/5000 dilution

    Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 2 : A431 (Human epidermoid carcinoma cell line) whole cell lysate
    Lane 3 : SK-BR-3 (Human mammary gland adenocarcinoma cell line) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 137 kDa
    Observed band size: 180 kDa why is the actual band size different from the predicted?



    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure time: Lane 1: 3 minutes; Lane 2: 3 seconds; Lane 3: 10 seconds.

    The molecular weight observed is consistent with the literature PMID :15741050.

  • All lanes : Anti-ErbB 2 antibody [EPR19547-12] (ab214275) at 1/1000 dilution

    Lane 1 : C6 (Rat glial tumor cell line) whole cell lysate
    Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 137 kDa
    Observed band size: 180 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The molecular weight is consist with literature PMID :15741050.

  • Immunohistochemical analysis of paraffin-embedded Human breast cancer tissue labeling ErbB 2 with ab214275 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Membrane staining on Human breast cancer is observed [PMID: 18437174].

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

  • Immunohistochemical analysis of paraffin-embedded Human breast tissue labeling ErbB 2 with ab214275 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Negative control: No staining on normal Human breast [PMID: 15150568].

    Counter stained with Hematoxylin.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed SK-BR-3 (Human mammary gland adenocarcinoma cell line) cells labeling ErbB 2 with ab214275 at 1/60 dilution (red) compared with a Rabbit IgG,monoclonal [EPR25A] - Isotype control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti Rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • ErbB 2 was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab214275 at 1/30 dilution.

    Western blot was performed from the immunoprecipitate using ab214275 at 1/1000 dilution.

    VeriBlot for IP secondary antibody (HRP) (ab131366), was used as secondary antibody at 1/1000 dilution.

    Lane 1: HeLa whole cell lysate 10µg (Input).

    Lane 2: ab214275 IP in HeLa whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab214275 in HeLa whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 3 seconds.

References

ab214275 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Mouse Derived Breast Cancer Cells)
Permeabilization
Yes - 0.5% Triton for 20 minutes
Specification
Mouse Derived Breast Cancer Cells
Blocking step
3% BSA + 5% Goat Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 26°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Nov 20 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (mammary epithelial)
Gel Running Conditions
Reduced Denaturing (10% gel)
Loading amount
40 µg
Specification
mammary epithelial
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Flavia Ghiraldini

Verified customer

Submitted Sep 15 2016

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Sign up